As shown in FIG. 1, human eye 100 is generally spherical and includes a tough outer layer, called the sclera 102, to which muscles (not shown) are attached which control the rotation of eye 100 within the eye socket. Beneath sclera 102 is a choroid layer 104, and beneath choroid layer 104 is a retina 106 which includes specialized photosensitive cells (rods and cones) that contain the visual pigment rhodopsin. Light enters eye 100 through a pupil 108 and is focussed onto retina 106 by a lens 110. An optic nerve 112 connects eye 100 to the brain.
Cone cells within the retina provide visual acuity and color vision, while rod cells are adapted for vision in dim light. Within rod and cone cells light sensitive molecules are located in a multilayered membrane structure. Within the multilayered membrane structure of rod cells, a linear polyene, 11-cis-retinal, is bound to a lysyl NH2 group of the membrane protein opsin via a Schiff's base. Within the multilayered membrane structure of cone cells, 11-cis-retinal is bound to membrane proteins that are similar to opsin. Light causes isomerization of 11-cis-retinal around the C(11)-C(12) double bond to form all-trans-retinal. This isomerization reaction triggers electrical changes in retina 106 which generate nerve impulses that are transmitted to the brain, via optic nerve 112, which interprets the nerve impulses to produce the sensation of sight.
The all-trans-retinal that is released from opsin, or other visual protein, is reduced to form all-trans-retinol (vitamin A) which is transported from the rod or cone cells into cells of the adjacent retinal pigment epithelium (RPE) where the all-trans-retinol is regenerated to 11-cis-retinal through a series of chemical reactions called the retinoid cycle. FIG. 2 shows a diagram of the retinoid cycle as it is presently understood. In brief, all-trans-retinol is esterified with fatty acids in a reaction catalyzed by lecithin:retinol acyltransferase (LRAT). All-trans-retinyl esters (referred to as retinyl esters) have been suggested to be the substrate for a putative isomerohydrolase in the RPE (Rando, R. R., “Membrane Phospholipids as an Energy Source in the Operation of the Visual Cycle,” Biochemistry 30:595-602, 1991) and for a retinyl ester hydrolase (REH) that produces all-trans-retinol, a substrate for the putative isomerase that converts all-trans-retinol to 11-cis-retinol which is then oxidized to form 11-cis-retinal. 11-cis-retinal is exported to the photoreceptors where it recombines with opsins to form 11-cis-retinylidene-opsins.
Defects in the retinoid cycle may lead to blinding disorders, for example, retinitis pigmentosa or macular degeneration. Among these, the inherited early onset dystrophies are collectively called Leber congenital amaurosis (LCA). LCA is a group of genetically heterogeneous diseases that involves, among others, mutations in the LRAT and RPE65 genes.
There is a continuing need for methods and devices that permit measurement of the levels of chemical components of the retinoid cycle, and changes in the levels of chemical components of the retinoid cycle, which can be used to assess the health of a mammalian retina, or can be used to assess a particular physiological state of a mammalian retina, or can be used to investigate, characterize, diagnose and/or treat a disorder of the mammalian retina.